This invention relates generally to a system and method for the preparation of arrays of separated molecules from a mixture of molecules.
Micro arrays having a matrix of positionally defined reagent target spots for performing chemical tests are known. Known reagents are deposited by spotting techniques well known in the art. In analyzing a sample, it is reacted with the array and separate chemical tests are performed with the reagent at each spot.
Mass spectrometers of various types have been used to identify molecules including proteins by mass analysis. The molecules are ionized and then introduced into the mass spectrometer for mass analysis. In recent years, mass spectrometers have been used by biochemists to identify both small and large molecules including proteins and to determine the molecular structure of the molecules including proteins.
Mixtures of biological compounds are normally separated by chromatographic techniques before the components of the mixture are mass analyzed. In some instances, chromatographically separated components of the mixture are used to create chips or arrays.
In proteomics the aim is to quantify the expression levels for the complete protein complement, the proteome, in a cell at any given time. The proteome is individual, environment and time dependent, and has an enormous dynamic range of concentration. Separation by two dimensional electrophoresis or electrophoresis and creation of spots on an array is cumbersome and slow. Modern analytical methods such as mass spectrometry are used for final analysis of the separated components of the protein complement.
Soft-landing of ions onto surfaces was proposed in 1977 [4] and successfully demonstrated two decades later [5]. Intact polyatomic ions were mass-selected in a mass spectrometer and deposited onto a surface at low kinetic energies (typically 5-10 eV). SIMS analysis was used to confirm the presence of a soft-landed species, C3H10Si2O35Cl+, on a fluorinated SAM surface. Evidence suggests that ions with sterically bulky groups have better deposition efficiencies than small ions [6]. Organic cations [7] and a 160-mer double-stranded DNA [8] (mass ca. 10 kDa) have also been soft-landed intact onto surfaces as have metal clusters [9]. In some of these cases there is evidence that the molecular entity on the surface is the ion, in others that it is the corresponding neutral molecule. There is even evidence [11] that intact viruses can be ionized, passed through a mass spectrometer under vacuum and collected and remain viable.
Separations are an inherent aspect of mass spectrometry; however, after mass separation of ions in vacuum, the resulting highly purified ionic materials are destroyed at the detector. Separations utilizing mass/charge ratios of gaseous ions have a different physical basis to those used in chromatography which are typically based upon chemical properties such as hydrophobicity, ionic strength in solution, and chemical functionality. This inherent difference indicates that mass spectrometric separation may be complementary to other forms of separation as a preparative technique, but previously developed mass spectrometry has not become a widespread method of collecting highly purified molecular species for preparation because of the overall inefficiency inherent in the ionization, ion transport, mass analysis, and ion collection processes.
The collection of materials from the mass spectrometer has been observed in a variety of experiments over several decades. Neutral species produced during electron impact of organic molecules [12] have been collected and characterized and used to examine mechanisms of fragmentation pathways. The collection of atomic ions as a result of collisions of mass-selected ions with surfaces is well-known from uranium isotope separations carried out during the Manhattan project. The collection of molecular ions at surfaces requires gentler conditions and ion soft landing was introduced for this purpose [13] (a variety of other phenomena [14] occur upon collision of molecular ions with surfaces, the best studied being surface induced dissociation [15-20]). Low kinetic energy (5-10 eV) beams of mass-selected organic ion beams can be collected at polyfluorinated self-assembled monolayer (SAM) surfaces [21]. Surface analysis confirmed the presence of deposited organic ion, such as the sterically bulky N,N-dimethylisothiocyanate cation. Organic cations [21,22], monomer cations to induce surface polymerization [24], polymers [25], and 160-mer double-stranded DNA [26] have also been soft-landed intact onto surfaces, as have metal clusters [27], and, in some experiments, virus particles have shown evidence of activity after in-vacuum deposition onto a collector plate following a gas-phase mass-selection [28]. In some of these cases there is evidence that the molecular entity on the surface is the selected ion [21], while in others it is the corresponding neutral molecule.
Growing demands from the proteomics and biotechnology fields for increased throughput of biological analyses have produced a rising interest in the production of protein microarrays from biological materials. Typically, fabrication of these arrays involves sample purification using various chromatographic techniques and micro-drop deposition methods [29-33]. The complementary separatory power of mass spectrometry to that of chromatography might allow experiments to address a broad range of problems in proteomics and related areas. Mass spectrometry using ion soft landing therefore appears to be a possible technology for the separation of biological compounds from mixtures and their storage in the array format for convenience of later analysis [34].
There is, therefore, a need for a new type of preparative mass spectrometer.